Identification of a regulatory cis-element within the 3′-untranslated region of the murine inducible nitric oxide synthase (iNOS) mRNA; interaction with heterogeneous nuclear ribonucleoproteins I and L and role in the iNOS gene expression

Söderberg, Malin; Raffalli-Mathieu, Françoise; Lang, Matti A.

doi:10.1016/j.molimm.2006.02.019

Cited by 29 publications

(18 citation statements)

References 40 publications

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“…The majority of the identified proteins are affiliated with mRNA translation. We chose to study PTB and hnRNP L because of the broad function of PTB as an ITAF (72) and because hnRNP L interacts with PTB (30,77). We found that Cat-1 mRNA associates with PTB and hnRNP L in the nucleus and continues to associate in the cytoplasm, consistent with the fact that PTB and hnRNP L are nuclear proteins that shuttle between the nucleus and the cytoplasm (42).…”

Section: Discussionmentioning

confidence: 72%

“…We propose that hnRNP L is a core factor in a multiprotein complex on the Cat-1 IRES that recruits the ribosome; depletion of hnRNP L during amino acid starvation limited binding of PTB to the IRES, but depletion of PTB did not affect binding of hnRNP L. Such cooperative action of hnRNP L and PTB was described for the lipopolysaccharide/gamma interferon-induced stability of inducible nitric oxide synthase mRNA stability through a mechanism involving PTB and hnRNP L binding to the 3Ј UTR (77). hnRNP L has been shown to have diverse functions (39).…”

Section: Discussionmentioning

confidence: 90%

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The hnRNA-Binding Proteins hnRNP L and PTB Are Required for Efficient Translation of the Cat-1 Arginine/Lysine Transporter mRNA during Amino Acid Starvation

Majumder

Yaman

Gaccioli

et al. 2009

Molecular and Cellular Biology

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The response to amino acid starvation involves the global decrease of protein synthesis and an increase in the translation of some mRNAs that contain an internal ribosome entry site (IRES). It was previously shown that translation of the mRNA for the arginine/lysine amino acid transporter Cat-1 increases during amino acid starvation via a mechanism that utilizes an IRES in the 5 untranslated region of the Cat-1 mRNA. It is shown here that polypyrimidine tract binding protein (PTB) and an hnRNA binding protein, heterogeneous nuclear ribonucleoprotein L (hnRNP L), promote the efficient translation of Cat-1 mRNA during amino acid starvation. Association of both proteins with Cat-1 mRNA increased during starvation with kinetics that paralleled that of IRES activation, although the levels and subcellular distribution of the proteins were unchanged. The sequence CUUUCU within the Cat Cationic amino acid transporter 1 (Cat-1) is a high-affinity Na ϩ -independent transporter of L-arginine and L-lysine belonging to system yϩ (12, 62). Growth factors, hormones, and nutrients can modulate its expression level (20,53). Expression of the Cat-1 gene increases during stress in a manner that involves phosphorylation of translation initiation factor 2␣ (eIF2␣) (24). During such conditions, expression of the Cat-1 gene is regulated at the level of (i) mRNA synthesis via the transcription factor ATF4, which binds an amino acid response element in the first exon of the gene (54); (ii) mRNA stability via the binding of the nucleocytoplasmic protein HuR to an AU-rich element present within its 3Ј untranslated region (UTR) (94); and (iii) translation via a cap-independent mechanism of initiation via an internal ribosome entry site (IRES) (23).IRES-dependent translation involves the recruitment of ribosomes independently of the m 7 G cap at the 5Ј end of the mRNA followed by initiation downstream of the ribosome binding site (46). We showed that increased translation of the Cat-1 mRNA during amino acid starvation requires the translation of a 48-amino-acid open reading frame (ORF) in the 5Ј UTR of the mRNA, introducing the concept of a "dynamic IRES" (23,93). In the absence of upstream ORF (uORF) translation, the Cat-1 IRES remains in an inactive conformation. During starvation, translation of the uORF unwinds this secondary structure, leading to formation of a conformation that has IRES activity. Previous studies have also proposed that the active conformation is stabilized by proteins (IRES transactivating factors [ITAFs]) that are either synthesized or modified by processes that require eIF2␣ phosphorylation. We also showed that decreased translation elongation rates of the uORF in fed cells result in a prolonged half-life of this active remodeled structure, mimicking the role of ITAFs in amino acid-starved cells (21). Ribosome stalling within the uORF can occur physiologically during stresses that cause increased phosphorylation of elongation factor 2 and therefore decreased translation elongation rates (63). These two mechanisms o...

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Section: Discussionmentioning

confidence: 72%

Section: Discussionmentioning

confidence: 90%

The hnRNA-Binding Proteins hnRNP L and PTB Are Required for Efficient Translation of the Cat-1 Arginine/Lysine Transporter mRNA during Amino Acid Starvation

Majumder

Yaman

Gaccioli

et al. 2009

Molecular and Cellular Biology

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“…However, recent evidence suggests that PTB is a major hnRNP protein with multiple roles in mRNA metabolism, including the stabilization of mRNAs like the CD154, insulin, and vascular endothelial growth factor mRNA (11)(12)(13)(14)(15). Moreover, PTB has been implicated in the regulation of the murine iNOS expression (27,37).…”

Section: Discussionmentioning

confidence: 99%

The Polypyrimidine Tract-binding Protein (PTB) Is Involved in the Post-transcriptional Regulation of Human Inducible Nitric Oxide Synthase Expression

Pautz¹,

Linker²,

Hubrich

et al. 2006

Journal of Biological Chemistry

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Human inducible nitric oxide synthase (iNOS) expression is regulated by transcriptional and post-transcriptional mechanisms. We have recently shown that the multifunctional RNAbinding proteins KH-type splicing regulatory protein and tristetraprolin are critically involved in the post-transcriptional regulation of human iNOS expression. Several reports have shown that KH-type splicing regulatory protein colocalizes with the polypyrimidine tract-binding protein (PTB), and both RNAbinding proteins seem to interact with the same mRNAs. Therefore we analyzed the involvement of PTB in human iNOS expression. In human DLD-1 cells, cytokine incubation necessary to induce iNOS expression did not change PTB localization or expression. However, intracellular binding of PTB to the human iNOS mRNA increased after cytokine stimulation. Overexpression of PTB resulted in enhanced cytokine-induced iNOS expression. Accordingly, small interfering RNA-mediated knock down of PTB reduced cytokine-dependent iNOS expression. Recombinant PTB displayed binding to an UC-rich sequence in the 3-untranslated region of the human iNOS mRNA. Transfection experiments showed that PTB mediates its effect on iNOS expression via binding to this region. The underlying mechanism is based on a modulation of iNOS mRNA stability. In summary, human iNOS is the first example of a human pro-inflammatory gene regulated by PTB on the level of mRNA stability.Post-transcriptional mechanisms represent an important part of the regulation of gene expression. In particular, genes whose expression has to be controlled precisely (e.g. oncogenes, pro-inflammatory genes) are regulated at the post-transcriptional level, mostly by modulation of mRNA stability (1, 2). Inherently unstable mRNAs that code for cytokines, transcription factors, proto-oncogenes, and pro-inflammatory mediators often contain AU-rich elements (AREs) 3 in their 3Ј-untranslated regions (3Ј-UTRs). These AREs are targets for transacting proteins regulating mRNA stability and translation (1, 3). The family of ARE-binding proteins includes the embryonic lethal abnormal vision protein family members (most importantly HuR), the ARE/poly(U)-binding/degradation factor 1 (AUF-1, also named hnRNP D), the KH-type splicing regulatory protein (KSRP), tristetraprolin, the T cell-restricted intracellular antigen (TIA-1), and the T cell-restricted intracellular antigen-related protein (1, 3). In mammalian cells, the AREsequences mediate mRNA decay mainly by recruitment of the exosome (a multisubunit particle with 3Ј to 5Ј nuclease activity) to the mRNAs, thereby promoting their rapid degradation. However, the mammalian exosome does not seem to recognize the ARE-containing RNAs on its own but requires certain AREbinding proteins (like KSRP or tristetraprolin) for this interaction (4, 5). The polypyrimidine tract-binding protein (PTB), also known as hnRNP I, is a major hnRNP protein with multiple roles in mRNA metabolism, including regulation of alternative splicing (6), internal ribosome entry site-driven translatio...

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“…Furthermore, an association of hnRNP L with the CA-rich cluster of the 3'UTR of VEGF confers stability to VEGF mRNA upon exposure to hypoxia [24]. However, deletion of the binding sites for hnRNP L in the 3'UTR of iNOS mRNA resulted in stabilization, indicating that hnRNP L promotes the degradation of iNOS mRNA [25]. We previously demonstrated that hnRNP L binds to CA repeats in 3'UTR of bcl-2 mRNA in vitro and in vivo [16].…”

Section: Discussionmentioning

confidence: 99%

“…In support of this conclusion, we previously observed that a larger complex with less mobility, in addition to the main complex, was formed with the riboprobe of CA repeats bcl-2 in REMSA (RNA electrophoretic mobility shift assays), both of which were supershifted with hnRNP L antibodies, indicating that the larger complex includes hnRNP L, CA repeats in bcl-2 mRNA and probably an unidentified protein [16]. Furthermore, hnRNP L has been shown to associate with several proteins such as AUF-1, hnRNP A2, hnRNP I and hnRNP L itself [17,25,26]. Thus, it is probable that interactions of hnRNP L with another protein via CA repeats of bcl-2 mRNA is an important prerequisite for the degradation of bcl-2 mRNA, which is not adequately disturbed by modulation of hnRNP L levels.…”

Section: Discussionmentioning

confidence: 99%

Effect of Modulation of hnRNP L Levels on the Decay of bcl-2 mRNA in MCF-7 Cells

Lim

Lee

Jung

et al. 2010

Korean J Physiol Pharmacol

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It has been shown that CA repeats in the 3'-untranslated region (UTR) of bcl-2 mRNA contribute the constitutive decay of bcl-2 mRNA and that hnRNP L (heterogenous nuclear ribonucleoprotein L) interacts with CA repeats in the 3'-UTR of bcl-2 mRNA, both in vitro and in vivo. The aim of this study was to determine whether the alteration of hnRNP L affects the stability of bcl-2 mRNA in vivo. Human breast carcinoma MCF-7 cells were transfected with hnRNP L-specific shRNA or hnRNP L-expressing vector to decrease or increase hnRNP L levels, respectively, followed by an actinomycin D chase. An RT-PCR analysis showed that the rate of degradation of endogenous bcl-2 mRNA was not affected by the decrease or increase in the hnRNP L levels. Furthermore, during apoptosis or autophagy, in which bcl-2 expression has been reported to decrease, no difference in the degradation of bcl-2 mRNA was observed between control and hnRNP L-knock down MCF-7 Cells. On the other hand, the levels of AUF-1 and nucleolin, transacting factors for ARE in the 3'UTR of bcl-2 mRNA, were not significantly affected by the decrease in hnRNP L, suggesting that a disturbance in the quantitative balance between these transacting factors is not likely to interfere with the effect of hnRNP L. Collectively, the findings indicate that the decay of bcl-2 mRNA does not appear to be directly controlled by hnRNP L in vivo.

show abstract

Identification of a regulatory cis-element within the 3′-untranslated region of the murine inducible nitric oxide synthase (iNOS) mRNA; interaction with heterogeneous nuclear ribonucleoproteins I and L and role in the iNOS gene expression

Cited by 29 publications

References 40 publications

The hnRNA-Binding Proteins hnRNP L and PTB Are Required for Efficient Translation of the Cat-1 Arginine/Lysine Transporter mRNA during Amino Acid Starvation

The hnRNA-Binding Proteins hnRNP L and PTB Are Required for Efficient Translation of the Cat-1 Arginine/Lysine Transporter mRNA during Amino Acid Starvation

The Polypyrimidine Tract-binding Protein (PTB) Is Involved in the Post-transcriptional Regulation of Human Inducible Nitric Oxide Synthase Expression

Effect of Modulation of hnRNP L Levels on the Decay of bcl-2 mRNA in MCF-7 Cells

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